7-hydroxy-cholesterol



Patented Nov. 16, 1937 v UNITED STATES PATENT OFFICE 7-HYDROXY-CHOLE STEROL Adolf Windaus, Goettingen,

and Friedrich Schenck, Wuppertal-Elberfeld, Germany, asslg'nors to Winthrop Chemical Company, Inc., New York, N. Y., a corporation of New York No Drawing. Application March 5, 1937, Serial No. 129,189. In Germany March 20, 1935 1 Claim.

10 This application refers to the said 7-hydroxycholesterol of the formular ncmcmcmon which is an intermediate. product in the said manufacture, and to its acyl derivatives. The '1- hydroxy-cholesterol forms white needle crystals which show a characteristic melting point of 5 178 C.; it is best characterized by its crystalline ester derivativesyits dibenzoate, for instance, is obtained in the form of white crystals which sharply melt at 17-25-4735 C. By saponification of the esters, the-'l-hvdroxy-cholesterol is recov- 30 ered. Our new 'l-hydroxy-cholesterol obtained from 'f-oxo-cholesterol, essentially difl'ers in many respects from the compound which is known in the literature under the name 0xy-,

cholesterin (compare, .for instance, Ber. der 35 deutschen Chem. Ges. 47, page'1454), hydroxycholesterol respectively, and which has been obtained from cholesterol by oxidation with benzoyl peroxide. The position of the second-hydroxyl group which is said to be introduced into the 40 cholesterol molecule by the said oxidation process is not known, as, on the whole, the chemical character of the saidnon-crystalline oxidationproduct is still to be elucidated. Contrary to 7- h'ydroxy-cholesterol the said hydroxy-cholesterol 45 is an amorphous light yellowsubstance, similar in appearance to amber. While the crystalline '1- hydroxy-cholesterol is difilcultly soluble in cold petroleum ether, the known hydroxy-cholesterol readily dissolves in cold petroleum ether. On

50 acting upon the known hydroxy-cholesterol with benzoyl chloride in the same manner as upon 'I- hydroxy-cholesterol no crystals are obtainable but only a varnish-like brilliant, glass-like transparent, absolutely amorphous substance (com 55 pare "Zeitschriit filr physiol. Chemie, 96, page acid anhydride.

345). In the so-called "Salkowskis test in which the sterol compound is dissolved in chloreform and concentrated sulfuric acid admixed thereto, our new 7-hydroxy-cholesterol yields a blue coloration of the chloroform layer and a red coloration with green fluorescence of the sulfuric acid layer, whereas the known hydroxy-cho lesterol yields a red coloration in the chloroform layer. The sulfuric acid layer also in this case has a red coloration with green fluorescence as is the case with most sterol compounds. Further-- more, a solution of 7-hydroxy-cholesterol in glacial acetic acid takes a pure blue coloration on the addition of concentrated sulfuric acid, whereas the known hydroxy-cholesterol under the same conditions yields a violet colored solution.

In accordance with our invention 7-hydroxycholesterol is obtained by reacting upon a 'l-oxocholesteryl-ester, for instance, 'I-oxo-cholesterylacetate, with an aluminium alcoholate, preferably with the alcoholate of a secondary alcohol, such as aluminium isopropylate in the presence of an alcohol. Advantageously the same alcohol is used as diluent which is present in the aluminium alcoholate. The treatment with the aluminium alcoholate has the effect that the 7-oxo group of the initial compound is reduced to the 'l-hydroxy group without simultaneously reducing the double bond in the 5.6-position of the cholesterol compound. But simultaneously with the reduction of the oxo group saponiflcation of the ester group takes place. The 'I-hydroxy-cholesterol formed is separated from the aluminium compound in the usual manner and is finally obtained by recrystallization in the form of white needles as indicated above. Its esters are obtained by the usual esteriflcationprocesses, for instance, by

means of benzoyl chloride, acetyl chloride, acetic The esteriflcation is advantageously carried out in the presence of a base, such as pyridine. On saponiflcation of the well crystallized esters of a sharp melting point, the 7- hydroxy-cholesterol is again obtained in the form of white needles which show a characteristic melting point of 178 'C'.

The invention is further illustrated by the following examples without being restricted thereto:

Example :For obtaining 'l-hydroxy-cholester- 01 50 grams of l-oxo-cholesteryl acetate are dissolved in 500 cos. of dry isopropyl alcohol and after the addition of 20 grams of liquefied aluminium isopropylate heated to boiling for 5 hours. After reduction is complete with the simultaneous splitting off of the acetyl group the isopropylalcoholic solution is diluted with ether. The

ethereal solution is first extracted with normal hydrochloric acid, then, after removing the aluminium hydroxide, with water. The dried ethereal solution is concentrated to about 100-150 ccs. and treated with the five-fold quantity of petroleum ether, whereupon abundant quantities of the 'Z-hydroxy-cholesterol separate in colorless jelly balls which are filtered with suction and washed with petroleum ether. By evaporating the mother lye to a small volume and by diluting with petroleum ether further quantities of 7- hydroxy-cholesterol may be obtained. on recrystallization the fl-hydroxy-cholesterol is obtained in the form of white needles, which on heating melt at about 173 C. The 7-hydroxycholesterol yields immediately an intensive blue coloration when heated on the water-bath with chloral hydrate. In the Salkowski reaction the sulfuric acid takes a red coloration, the chloroform a blue coloration, an intensive green fluorescence occurring simultaneously. With antimony' trichloride in chloroform a deep bluecoloration is obtained after a short time.

For obtaining 7-hydroxy-cholesteroldibenzoate grams of '7-hyclroxy-cholesterol are dissolved in 200 ccs. oi pyridine and treated with a mixture 01' 30 grams of. benzoylchloride and 30 grams of pyridine. After standing for 24 hours, the ben- -zoyiation product is precipitated with 600ccs. of

Patent N 0. 2,098,985.

ADOLF 'WINDAUS ET AL.

It is hereby certified that error shaken with 500 cos. of fresh water. The water is poured oil and "the crude benzoate, covered with methyl alcohol, is left standing for several hours,

' whereupon it gradually turns to a white crystal powder.

The latter is filtered, washed with methyl alcohol and recrystallized from ethermethyl alcohol foriurther purification. In this manner thin needles are obtained in a good yield which melt at 1725-1735 C.- The 7-hydroxycholesterol-dibe'nzoate displays the same color reactions as the l-hydroxy-cholesterol. By saponification the 7-hydroxy-cholesteroi is recovered in the form of crystals showing the properties indicated above. a

In an analogous manner the 7-hydroxy-cholesterol-diacetate is obtained when reacting upon 10 grams of 7-hydroxy-cholesterol with 10 grams of acetic acid anhydride in the presence of cos. of pyridine. The diacetate exists in two isomorphous forms which melt at 98 C.100 C. and 106.5 C.- 107.5 C. respectively.

We claim:-

7-hydroxy-cholesterol, forming white crystals yielding an intensive blue coloration when heated on the water-bath with chloral hydrate, yielding in the Salkowski reaction a blue coloration of the chloroform solution, forming in the form of its dibenzoate white crystals melting at 1725-1735 C.

WINDAUS. FRIEDRICI-I SCHENCK.

I Certificate of Correction November 16, 1937.

ppears in the printed specification of the above numbered patent requiring correction as follows: Page 1, first column, line 15, in

the formula, for

read

I CH

CH.CH OH:CHgCH v U CHi and that the said Letters Patent should be read with this correction therein that the same may conform to the record of the case in the Patent Oifice.

S1gned and sealed this 8th day of February, A. i938.

[SEAL] 'HENRY" VAN ARSDALE,

ethereal solution is first extracted with normal hydrochloric acid, then, after removing the aluminium hydroxide, with water. The dried ethereal solution is concentrated to about 100-150 ccs. and treated with the five-fold quantity of petroleum ether, whereupon abundant quantities of the 'Z-hydroxy-cholesterol separate in colorless jelly balls which are filtered with suction and washed with petroleum ether. By evaporating the mother lye to a small volume and by diluting with petroleum ether further quantities of 7- hydroxy-cholesterol may be obtained. on recrystallization the fl-hydroxy-cholesterol is obtained in the form of white needles, which on heating melt at about 173 C. The 7-hydroxycholesterol yields immediately an intensive blue coloration when heated on the water-bath with chloral hydrate. In the Salkowski reaction the sulfuric acid takes a red coloration, the chloroform a blue coloration, an intensive green fluorescence occurring simultaneously. With antimony' trichloride in chloroform a deep bluecoloration is obtained after a short time.

For obtaining 7-hydroxy-cholesteroldibenzoate grams of '7-hyclroxy-cholesterol are dissolved in 200 ccs. oi pyridine and treated with a mixture 01' 30 grams of. benzoylchloride and 30 grams of pyridine. After standing for 24 hours, the ben- -zoyiation product is precipitated with 600ccs. of

Patent N 0. 2,098,985.

ADOLF 'WINDAUS ET AL.

It is hereby certified that error shaken with 500 cos. of fresh water. The water is poured oil and "the crude benzoate, covered with methyl alcohol, is left standing for several hours,

' whereupon it gradually turns to a white crystal powder.

The latter is filtered, washed with methyl alcohol and recrystallized from ethermethyl alcohol foriurther purification. In this manner thin needles are obtained in a good yield which melt at 1725-1735 C.- The 7-hydroxycholesterol-dibe'nzoate displays the same color reactions as the l-hydroxy-cholesterol. By saponification the 7-hydroxy-cholesteroi is recovered in the form of crystals showing the properties indicated above. a

In an analogous manner the 7-hydroxy-cholesterol-diacetate is obtained when reacting upon 10 grams of 7-hydroxy-cholesterol with 10 grams of acetic acid anhydride in the presence of cos. of pyridine. The diacetate exists in two isomorphous forms which melt at 98 C.100 C. and 106.5 C.- 107.5 C. respectively.

We claim:-

7-hydroxy-cholesterol, forming white crystals yielding an intensive blue coloration when heated on the water-bath with chloral hydrate, yielding in the Salkowski reaction a blue coloration of the chloroform solution, forming in the form of its dibenzoate white crystals melting at 1725-1735 C.

WINDAUS. FRIEDRICI-I SCHENCK.

I Certificate of Correction November 16, 1937.

ppears in the printed specification of the above numbered patent requiring correction as follows: Page 1, first column, line 15, in

the formula, for

read

I CH

CH.CH OH:CHgCH v U CHi and that the said Letters Patent should be read with this correction therein that the same may conform to the record of the case in the Patent Oifice.

S1gned and sealed this 8th day of February, A. i938.

[SEAL] 'HENRY" VAN ARSDALE, 

